We have been studying three regulatory elements of the human c-myc protooncogene and the proteins which bind to them. 1) 1.5 kb upstream of promoter P1 resides a cell-type and differentiation specific positive cis-element. A novel protein binding to this element has been purified and subjected to proteolytic degradation and partial sequence determination. The information thus obtained allowed the cloning of a gene encoding a novel DNA binding protein. This protein possesses a structure comprised of alternating amphiphathic helices (five) and repeating units (four). The minimal sequence specific DNA binding domain is composed of two helices and two repeats. Analysis of the expression of this factor reveal it to be tissue specific and highly regulated. Functional characterization of the protein is ongoing. This protein binds specifically to the FUSE element in either single or double-stranded form. 2) 100-150 bp upstream of P1 a complex set of trans-factors binds to a cytidine-rich element repeated five times. Some of the factors which interact with this element possess the ability to recognize specific single-stranded sequences. One of the pyrimidine-rich strand binding proteins is hnRNP protein K. In vitro and in vivo evidence strongly implicates hnRNP protein K as a trans-activator of this site. Additional proteins interact with the purine strand. The purine strand binding factors possess several unusual properties which correlate well with the activity of this element in in vitro transcription systems. One of these factors has been identified as an altered form of a zinc finger protein. The purification of the remaining factors with an aim of cloning their genes is in progress. 3) Approximately 1 kb downstream of c-myc promoter P1 is an element which has been found to be mutated frequently in Burkitt lymphoma. Several proteins have been shown by cross-linking or binding with SDS-PAGE purified and renatured polypeptides to interact with this element. Some of these proteins possess unusual properties which suggest interesting regulatory mechanisms for controlling c-myc expression. Based upon newly derived peptide sequence, the identification of one of these factors as a new member of a known family of DNA binding proteins has been made. Cloning of this factor is in progress.